Mechanisms governing steroidogenesis and the development and differentiation of steroidogenic organs, namely the adrenal glands and gonads, remain a fundamental question in mammalian biology. A functional adrenal cortex is essential for life as evidenced by the early death of patients with congenital adrenal hypoplasia or adrenal steroid deficiencies. Recently, the orphan nuclear receptor Steroidogenic factor-1 (SF-1) has been shown to be essential for both gonadal and adrenal development, as evidenced in SF-1 null mice created by gene targeted disruption of the FTZ-F1 gene. These mice lack adrenal glands, gonads and the gonadotrope cell type of the pituitary gland. Our laboratory has been studying the molecular biology of SF-1 and its role in gonadal development. SF-1 is a member of the monomer-binding subfamily of orphan nuclear receptors. What remains to be defined for this subfamily are the mechanisms by which they activate or repress their target genes. Specifically, it is critical to know if these orphans, including SF-1, are activated by ligand and/or post-translational modification/ Furthermore, their activation may involve the active translocation of receptor to the nucleus prior to activation of gene transcription. The inability to demonstrate convincing transactivation of target genes by SF-1 suggests that for SF-1 a cofactor or ligand is required. Our laboratory is now able to reconstitute fully activated SF-1 in a cell culture system by addition of semi-purified testis extract. This system enables me to now study the precise mechanisms of transactivation by a prototypic member of the monomer-binding orphan nuclear receptor subfamily. The phenotype created in the SF-1 null ice is reminiscent of the clinical syndromes of congenital adrenal hypoplasia, where the adrenal glands are hy - or aplastic and the patients are often hypogonadal. Conversely, in congenital primary adrenal hyperplasia, the adrenal glands are hyper- or dysplastic and patients are at times hypergonadal with gonadal steroid cell neoplasms. Only one gene associated with the X-linked form of congenital adrenal hypoplasia has been cloned. This gene has been found to code another orphan nuclear receptor, DAX-1. This leaves open the possibility that the other syndromes of primary adrenal hypoplasia and hyperplasia are linked directly to the nuclear receptors, SF-1 and DAX-1. In this application, I propose to examine the molecular mechanisms of SF-1 gene activation and begin to characterize the role of SF-1 in adrenal cortical cell lineage determination.